The invention relates to a miniature high-frequency ventilator for small animals or human infants.
Genetically engineered mice, made possible by techniques of molecular biology, are a valuable resource in furthering both the medical and biological sciences. By changing the chromosomal content in specific ways, various proteins can be knocked out or altered, whose expression or lack thereof can be investigated. The mice are a choice animal for such studies. Presently, much of their genome has been mapped. Additionally, like humans, they are mammals and therefore share many homologous chromosomes. By studying mice, further insight into human function can be gained. Furthermore, the mice can be bred with regularity and ease.
Unfortunately, the physical expression of mutations can sometimes be detrimental to the well-being of the mice. Various problems such as cardiac or respiratory failure can occur, causing premature death. Presently, there is no equipment designed to sustain newborn animals of this size, and such problems go unsolved.
The need for a miniature mechanical ventilator for newborn mice has arisen from recent studies on NMDA-R1 genetically engineered mice. The NMDA-R1 molecule is a key component of NMDA receptors. These receptors are believed to play a critical role in the plasticity of the nervous system. Unfortunately, the mice die within the first 10 to 20 hours after birth--prior to most major neurological developments--precluding further studies into the role of NMDA receptors in learning and memory.
The recessive mutation has various side effects. Some studied abnormalities include loss of hind leg motor control and balance, improper mastication (leading to an inability to suckle milk), and a degradation of the respiratory system. Observation of these symptoms has proven to be an effective determinant of the genetic make up (either homozygous recessive, or homozygous/heterozygous dominant) when compared to genetic prototyping data.
Previous studies have shown that the respiratory failure is preceded by periods of apnea and respiratory instability. FIGS. 1A and 1B respectively show graphical representations of breathing samples for normal and mutant newborn mice. One hypothesis that has been raised is that the early deaths of the newborn mutants are caused by this failure.
By artificially ventilating the "knockout" mice, the respiratory system may be preserved, prolonging the lives of the mice for a more thorough study into the roles of the NMDA receptors in neuroplasticity and neural development.
Current animal ventilators are not suitable to ventilate neonatal mice. They are designed to deliver positive pressure at the airways via pistons, diaphragms, or pressure sources. Being designed for larger animals, the devices generally deliver volumes of at least a few milliliters. Previous studies have shown that the newborn mice have tidal volumes ranging from 0.015 to 0.045 ml. Existing devices would therefore be imprecise over the required range. Such unreliability could result in uncontrolled volume generation and barotrauma for the fragile newborns.
A further problem with the current ventilators is that creating a positive pressure at the airway generally requires an intratracheal connection. Though this surgical procedure can be performed with some confidence in larger animals, a tracheotomy is too difficult to perform on newborn mice. Generally, this technique requires opening the throat and inserting a tube into the thrachea, an extremely invasive operation considering the size and fragility of the neonates. Such trauma makes the use of this ventilatory style an impossibility in the mutant mice.
A final concern with current methods of animal mechanical respiration is that the ventilators deliver automatic breaths at regular intervals. Such mechanically controlled breathing creates a problem, especially since the knockout mice breathe over a wide range of frequencies. The normal frequency is about 120 breaths per minute (bpm). However, depending on the stage of respiratory distress, much lower frequencies are observed (60 bpm). Delivering automatic breaths at a significantly different frequency than the spontaneous rate can be uncomfortable, and moreover, generate ventilator fighting. This condition arises when some of the animal breaths are out of phase with the mechanical breaths (i.e. the animal expires while the ventilator gives a positive inspiratory pressure). The competing efforts result in inefficiency and can cause further respiratory trauma to the already sick animal.
To minimize the invasiveness of the subject/ventilator connection, and reduce the risk of barotrauma, a negative pressure strategy can be employed. Instead of creating positive airway pressure, a negative pressure is generated around the body surface as shown in FIGS. 2A and 2B. FIGS. 2A and 2B respectively show diagrams of methods of ventilation with positive and negative pressure. Such techniques were used on humans in the 1950s and 60's with devices aptly named "iron-lungs." These machines were bulky and inefficient, and therefore quickly outmoded with the advent of positive pressure respirators. However, studies have shown thoracic oscillations to be as effective as tracheal methods in allowing blood gas exchange. An advantage that the negative pressure technique has in newborn mouse ventilation is that the only subject connection is an external cuff around the neck to isolate the body from atmospheric pressure.
Two methods to overcome the problem of ventilator fighting are through patient triggered systems, such as pressure support and negative impedance ventilation, or through high frequency ventilation (HFV).
For human ventilation, patient triggering systems have been developed, where the breathing effort of a patient can be sensed via pressure of flow fluctuations. The effort is then assisted with an inspiratory pressure. Unfortunately, the pressures and flows generated by a newborn mouse are too minute to be sensed reliably. A high threshold would mean a significant time delay in respiratory assistance, while a low threshold would result in excess breathing due to noise.
High frequency ventilation considers an alternate method of breathing by fluctuating respiratory pressures at several times the normal frequency in the range of 10 to 30 Hz. To prevent over-ventilation and hypocapnia, the tidal volume is accordingly reduced by an order of magnitude. This process results in simply oscillating the air within the lungs. Previous studies performed on larger animals have revealed that HFV provides adequate gas exchange in the alveoli, and is therefore an effective method of ventilation.
With these considerations, a miniature HF negative pressure ventilator in accordance with the present invention is presented. However, the small size and fragility of the newborn mice creates a more complex problem. Since minute volumes are generated, the leakage in the system must be minimized in a non-invasive manner. Also, previous systems applied both a positive and negative pressure to the body surface, with a mean pressure equal to atmospheric. These devices used on rats have proven to be an effective means of ventilation. Although such pressure fluctuations are acceptable in small volume generation, if larger volumes are needed, the positive surface pressure could pose a serious threat to the subject by closing the airways.
Since mechanical respiratory attempts have never been made on newborn mice, a large specification range was needed. The ventilator of the present invention is designed to deliver volumes from 0.00 ml to 0.04 ml over a frequency band of 1 Hz (in the normal range) to 100 Hz (higher than the present limits of HFV).